Biochemische und biophysikalische Charakterisierung des GARP2-Proteins aus Sehstäbchen der Rindernetzhaut

Abarca Heidemann, K.

Dissertation / PhD Thesis/Book

Biochemische und biophysikalische Charakterisierung des GARP2-Proteins aus Sehstäbchen der Rindernetzhaut

Abarca Heidemann, K. (Corresponding author)FZJ*

2002
Forschungszentrum Jülich Zentralbibliothek, Verlag Jülich

Jülich : Forschungszentrum Jülich Zentralbibliothek, Verlag, Berichte des Forschungszentrums Jülich 4061, IX, 130 p. (2002) = Köln, Univ., Diss., 2002

Please use a persistent id in citations: http://hdl.handle.net/2128/2568

Report No.: Juel-4061

Abstract: The GARP proteins of rods have been localized exclusively at the disc incisures and rims that are in close proximity to the plasma membrane. GARP2 is the most abundant GARP species, and a major protein in ROS. The localization and the amount of GARP2 indicate that it may have an important role in rod phototransduction. Studies focusing on the protein/protein interactions involving GARP proteins have revealed contradictory results (Körschen et al., 1999 ; Poetsch et al., 2001). The present work describes the purification of native GARP2 from bovine ROS. Furthermore this work describes the cloning, expression and purification of two different recombinant GARP2 proteins. A detailed analysis of the structural and biochemical characteristics of the purified proteins shows that the unusual running behavior of GARP2 is not due to dimerisation. The results also strongly suggest that GARP2 is not a globular protein. Moreover the GARP2 secondary structure was analyzed by CD. The interaction of GARP2 with proteins was studied by gel filtration, affinity chromatography, and crosslinking. These experiment show that GARP2 interacts with RDS/peripherin and possibly also with RGS9. Besides, an interaction to unknown proteins with an apparent M$_{w}$ of $\sim$18 kDa, $\sim$ 30 kDa and $\sim$ 70 kDa could be detected. The interactions with the PDE, the GC, and the ABCR could not be proven. Using the purified GARP2 proteins, the effect on the activated PDE was examined. The previously shown powerful inhibition of PDE by the recombinant HT-GARP2 could be reproduced, but using native GARP2 no inhibition was observed. These results suggest that the fusion part of the HT-GARP2 is responsible for the inhibitory effect. These findings and the monoclonal antibodies against the different GARP proteins, provide a solid basis to perform further biochemical, immunohistochemical and cristallographic studies.

Note: Record converted from VDB: 12.11.2012
Note: Köln, Univ., Diss., 2002

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